Revisiting the HO●-initiated oxidation of L-proline amino acid in the aqueous phase: influence of transition metal ions

The oxidation of L-proline (Pro) by HO● radical in water and the influence of transition metal ions on this process has been revisited by using the density functional theory (DFT) method at the M05-2X/6-311 + + G(3df,3pd)//M05-2X/6-311 + + G(d,p) level of theory at the temperature of 298.15 K. The main reactive sites of the HO●–initiated oxidation of Pro via hydrogen atom transfer (HAT) reactions are at the β- and γ-carbon, with the branching ratios being 44.6% and 39.5%, respectively. The overall rate constant at 298.15 K is 6.04 × 108 M−1 s−1. In addition, Pro tends to form stable complexes with both Fe and Cu ions via the –COO functional group of dipole-salt form. The most stable Cu(II)-Pro complexes have high oxidant risks in enhancing the HO● formation in the presence of reducing agents. Besides this, the high oxidation state metal complexes, i.e. Fe(III)-Pro and Cu(II)-Pro, may be oxidized by HO● radical via HAT reactions but with a lower rate constant than that of free-Pro. By contrast, the low oxidation state metal complexes (i.e. Fe(II)-Pro and Cu(I)-Pro) have higher oxidation risks than the free ligands, and thus, the complexation enhances the oxidation of Pro amino acid.

plus selenocysteine, appearing in the genetic code [2]. In the biological environment, the amino acid consists of monomers for several biological macromolecular structures such as proteins, neurotransmitters, hormones and nucleic acids [3].
Several factors, such as free radicals and ultraviolet radiation, may oxidize the amino acids in different environments [6][7][8][9][10][11][12][13][14]. Hydroxyl radical (HO • ) produced by the Fenton reagent, i.e. -Fe(II) + H 2 O 2 system, consists of one of the most reactive oxidative factors investigated by Stadtman et al. [6]. The obtained products of the oxidation of amino acids by HO • are NH þ 4 , α-ketoacids, CO 2 , oximes and aldehydes or carboxylic acids containing one less carbon atom. Furthermore, the relative oxidation rates of the studied amino acids varied as follows: L-leucine (Leu) > L-serine (Ser) > L-alanine (Ala) > glycine (Gly) > L-arginine (Arg) > L-tryptophan (Trp) > L-methionine (Met) > L-asparagine (Asp) > L-histidine (His). Besides, McGregor et al. [7] studied the reactions between singlet oxygen, O 2 ( 1 Δ g ), with four amino acids, including His, Met, Trp and L-tyrosine (Tyr). As a result, the reaction rates varied from 5 × 10 5 to 6 × 10 7 M −1 s −1 , with the decreasing order being: His > Trp > Tyr > Met. The influences of temperature and pH on the oxidation of Gly, Ala, Ser and L-threonine (Thr) in the aqueous phase by HO • radical are experimentally and theoretically performed [8]. The rate constant increases from 1.3 to 3.0 times with a temperature varying from 278 to 318 K and from 1.2 to 3.6 times with a pH of 1.0 to 6.0.
Formation of complexes between amino acids and transition metal ions, such as Fe(III)/Fe(II) and Cu(II)/Cu(I), plays essential roles in many metabolisms in living creatures, i.e. electron transfer, catalysis, structural support and protein folding/unfolding [15][16][17][18][19][20]. However, these metal complexes may enhance the production of reactive HO • radicals [21,22]. Milach et al. [21] found that L-glutamic acid (Glu), Ala, His and L-cysteine (Cys) and its derivatives favour the HO • formation in the presence of cupric ion -Cu(II). Furthermore, Cys also enhances the HO • production in the Fe(II)/EDTA system. Truong et al. [23] theoretically showed that Leu has high chelating potential toward Fe and Cu ions via its -COO functional group of dipole-salt form. The most stable Cu(II)-Leu complexes significantly promote the HO • radical formation via Fenton reactions. Remarkably, the oxidant activity of complexes is enhanced considerably in the presence of an ascorbate anion.
L-(-)-Proline (Pro), a cyclic α-amino acid, is the only one of 20 DNA-amino acids having a secondary α-amine group (figure 1) [24]. Humans can produce it from arginine (Arg) and glutamine/glutamate (Gln) [24]. Pro is one of the main collagen components in the skin, tendons, bones and connective tissue, promoting their health and healing abilities. Pro plays a role as a biological antioxidant as reactive oxygen species (ROS) scavenger and singlet oxygen quencher [25,26]. Pro also prevents lipid peroxidation in alga cells exposed to heavy metals [27]. Direct oxidation of amino acid side chains of Pro, Arg, and L-lysine (Lys) may lead to the formation of glutamic semialdehyde and αaminoadipicsemialdehyde, respectively, which are the protein carbonyl derivatives [28]. It is noted that high levels of protein carbonyls resulting from protein oxidation are implicated in either ageing [28,29] or the aetiology, progression or manifestations of several diseases such as rheumatoid arthritis [30], muscular dystrophy [31], and Alzheimer's disease [32]. Because of the direct relationship between protein oxidation and different diseases, several attempts in the literature have contributed to a better understanding of the Pro oxidation by reactive oxygen species (ROS) and other amino acids involved in the proteins. For example, Salamone et al. measured the rate constant for the HAT reaction between the cumyloxyl radical -[PheC(CH 3 ) 2 ]O • with N-boc-protected Pro amino acid and its dipeptides [33]. The total rate constant of the reaction found in DMSO solvent at 25°C is (3.3 ± 0.2) × 10 6 M −1 s −1 . The authors suggested that the HAT reaction at the δ-C−H bond (i.e. C7 position, royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 10: 230114 figure 1) is the predominant one due to the electron-withdrawing character of the α-substituents of the C−H bonds closer to the amino acid backbone. This polar effect deactivates the HAT reaction to electrophilic radicals such as HO • or Cl • . Conversely, Signorelli et al. computationally indicated that the HO • -initiated oxidation of Pro occurs more favourably at the βand γ-carbon positions (i.e. C5 and C6, figure 1) than those at the αand δ-carbons (i.e. C4 and C7, figure 1) with lower reaction free energies of about 2 kcal mol −1 [34].
Although many works have focused on Pro's oxidation activity, its mechanism remains contrary and needs more attempts to shed light on the reaction kinetics and different influencing factors, such as temperature and transition metal ions. Thus, in this work, we revisit the HO • -initiated oxidation of Pro at the temperature of 298.15 K. For the first time, we investigate the effect of transition metal ions on the oxidation mechanism. First, we study the kinetics of the oxidation reaction by HO • radical via formal hydrogen transfer (FHT) and single electron transfer (SET) mechanism. Second, the indirect effect of the complexation between Pro and Fe/Cu ions on the HO • production is examined by evaluating the reduction reactions of the complexes towards two reducing agents, i.e. superoxide radical anion (O 2 •-) and ascorbate anion (Asc -), which are available in human body environment.
Finally, the reactions between the most stable Fe-Pro and Cu-Pro complexes with HO • radical are investigated to examine the direct effect in the oxidation process resulting from the metal ions' chelation.

Computational method
All geometry optimization and vibrational frequency calculations were performed by Gaussian 16 Rev.
A.03 package [35]. All structures of reactants, transition states, pre-reactive complexes, post-reactive complexes and products were studied in the aqueous phase using M05-2X functional [36] and 6-311 + + G(d,p) basis set. Single-point calculations then improved the accuracy of the energy values at the M05-2X/6-311 + + G(3df,3pd) level of theory. The zero-point energy (ZPE) was corrected using a scaling factor of 0.961 [37,38]. The M05-2X functional has largely been chosen by several similar kinetic studies related to the complexes of transition metal and an organic ligand [23,[39][40][41]. The influence of the aqueous media was mimicked using the solvation model based on the quantum mechanical charge density of a solute molecule interacting with a continuum description of the solvent (SMD) [42][43][44], which has also been widely used in the literature [39,41].  4 ] + were employed as a recommendation by previous works [40,41,[45][46][47]. The high-spin states of hydrated Fe(II) and Fe(III) ions in this study were equal to 5 and 6, respectively [23,41,46,[48][49][50][51]. The kinetics of HO-initiated oxidation of Pro via FHT and SET reaction, the energetics for the chelation processes of Pro toward copper and iron ions, and the pro-oxidant risks of Pro determined via the reduction reactions of Fe(III)-to-Fe(II) and Cu(II)-to-Cu(I) complexes by two reducing agents, the superoxide radical anion (O 2

•-
) and the ascorbate anion (Asc − ) were resumed in the electronic supplementary material (SI file).

Results and discussion
3.1. The oxidation of L-proline by HO • radical Optimized geometry for neutral and dipole salt forms of Pro in the aqueous phase are shown in figure 1. The frontier orbitals distribution and electrostatic potential maps are shown in electronic supplementary material, figure S1, and its Cartesian coordinates are shown in electronic supplementary material, table S1 (ESI). It is noted that the dipole-salt form is the primary form of Pro in the aqueous phase (approx. 100%). Thus, the overall rate constant of its oxidation reaction will be considered based on the dipole-salt form.
In this section, the oxidation of Pro by HO • radical is estimated by investigating the kinetics of the reactions occurring via two processes, formal hydrogen transfer (FHT) and single electron transfer (SET).  . It is noteworthy that the Δ r G 0 of FHT reactions are negative, from −25.2 to −20.0 kcal mol −1 ; therefore, these reactions are, as expected, favourable and spontaneous. The most negative Δ r G 0 value is found at the H9 position, being −25.2 kcal mol −1 . By contrast, the Δ r G 0 of SET reaction is highly positive, being 27.1 kcal mol −1 . Regarding the Gibbs free energies of activation, the ΔG ‡ values for FHT reactions of the dipole-salt form stand from 5.3 to 12.3 kcal mol −1 . Conversely, the ΔG ‡ of SET reaction is much higher, i.e. 86.5 kcal mol −1 .
Furthermore, the diffusion-corrected apparent rate constants of FHT reactions for the dipole-salt form vary from 5.03 × 10 2 to 1.81 × 10 8 M −1 s −1 , observed at the H16 (Γ 0.0%) and H10 sites (Γ 30.0%), respectively (figure 2). The reaction occurring at the H12 position also attracts our attention, with the rate constant as high as that at H10, i.e. 1.55 × 10 8 M −1 s −1 (Γ 25.6%). The total branching ratios for the FHT reactions at H10/H11 (of C5 carbon) and H12/H13 (of C6 carbon), 44.6% and 39.5%, are remarkably higher than that at H9 (of C4 carbon, 11.5%) and H14/H15 (C7 carbon) (i.e. 4.2%). Our results agree with those computationally observed by Signorelli et al. [34], which indicated that the HO • -initiated oxidation of Pro in the aqueous phase occurs more favourably at the βand γ-carbon positions than those at the αand δ-carbon. Our observation is also in accordance with the ones found in previous studies. Galano et al. found that HO • oxidizes Asp more favourably at the β-carbon (k, 2.66 × 10 9 L mol −1 s −1 ) than at the α-carbon (k, 2.41 × 10 6 L mol −1 s −1 ) at 298.15 K in the gas phase [56]. In the same reaction conditions, the authors observed that the HO • -initiated oxidation of Met via HAT reaction is more dominant at the γ-carbon (k, 1.66 × 10 10 L mol −1 s −1 ) than at the α-carbon (k, 7.95 × 10 6 L mol −1 s −1 ) [57]. Uranga et al. also confirmed that the first HO • abstracts an H-atom from the β-carbon atom in the case of Ser and Thr, whereas γ-carbon and γ-sulfur atoms are the preferred sites for Met and Cys, respectively [12]. Alternatively, the SET reaction of the dipole-salt form of Pro is insignificant, with a negligible branching ratio. royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 10: 230114 In conclusion, the overall rate constant of the oxidation reactions is equal to 6.04 × 10 8 M −1 s −1 at 298.15 K, which is close to the diffusion limit. This value is about 100 times higher than that obtained with N-boc-protected Pro oxidized by the cumyloxyl radical ([PhC(CH 3 ) 2 ]O • ) in DMSO solvent at 25°C, with the value being (3.3 ± 0.2) × 10 6 M −1 s −1 [33]. The obtained results can be explained by the fact that Pro exists in DMSO in both the neutral (30%) and dipole-salt forms (70%), which is quite different from our observation in water (approx. 100% dipole-salt form), and by the lower reactivity of the cumyloxyl radical compared to the hydroxyl one. Furthermore, this rate constant is significantly higher than the HO • -initiated oxidation of His, Met, Trp, Tyr and Ala [8] but lower than those of Asp [56], Ser [58] and Leu [59].

The complexation activity of L-proline towards transition metal ions
The complexation reactions of Pro were investigated with the ferric/ferrous and cupric/cuprous ions to shed light on the possibility of forming stable complexes between Pro and the transition metal ions. We evaluated the complexes of both neutral (Neu) and dipole-salt (DpS) forms. The neutral form of Pro has two chelating sites, including O2-Neu, and Site-1, while the dipole-salt has three chelating positions: O1-DpS, O2-DpS and Site-2 (figure 1). For the mono-ligand complex, the chelation sites may be O2-Neu, O1-DpS and O2-DpS for monodentate complexes and Site-1 and Site-2 for bidentate complexes. The chelation sites for the di-ligand complexes having two Pro ligands are only found at Site-2. In this section, we only resume the results obtained with the dipole-salt form because of its predominance in the water.  Table 1. The Gibbs free energies (Δ r G 0 , kcal mol −1 ), Gibbs free energies of activation (ΔG ‡ , kcal mol −1 ), diffusion rate constants (k D , M −1 s −1 ), thermal rate constants (k, M −1 s −1 ), apparent rate constants (k app , M −1 s −1 ) and branching ratios (Γ, %) of the FHT and SET reactions between the Pro-dipole-salt form with HO • radical calculated in water at 298.15 K. Gibbs free energies (Δ r G 0 ) and formation constants (K f ) obtained with the Pro-dipole-salt are resumed in table 2. The complete data of all Fe(III)/Fe(II)-Pro complexes are shown in electronic supplementary material, table S5 (ESI). As seen in table 2, the formation of all the Fe(III)-Pro and Fe(II)-Pro complexes in dipole-salt form is favourable and spontaneous, with the negative Δ r G 0 values varying from −9.1 to −5.8 kcal mol −1 and −8.7 to −3.4 kcal mol −1 , respectively. Their formation constant K f is remarkably high, ranging from 1.71 × 10 4 to 4.52 × 10 6 and 3.33 × 10 2 to 2.42 × 10 6 , in turn. Site-2 is the preferable chelating site for both Fe(III)-Pro and Fe(II)-Pro complexes. The di-ligand complex at Site-2 demonstrates the chelating ability of Pro at its higher concentration. We can see that the complexation  Table 2. Gibbs free energies (Δ r G 0 , kcal mol −1 ) and formation constants (K f ) of the complexation reactions between Pro-dipolesalt forms and Fe/Cu ions in the aqueous phase. Calculation are performed at M05-2X/6-311 + + G(3df,3pd)//M05-2X/6-311 + + G(d,p) level of theory.

. Indirect enhancement
In this section, we try to address whether the complexation of Pro with transition metal ions, like Fe(III) and Cu(II), affects its oxidation. Indeed, several previous studies showed that some polyphenol-based molecules indirectly exhibit the reactive HO • radical production via the Fenton-like reactions of their transition metal ion complexes [23,40,41,[45][46][47]. The formed HO • radicals have a high potential to return oxidizing Pro leading to the pro-oxidant risk of an organic molecule.
Thus, we evaluate the reduction reactions of Fe(III)-to-Fe(II) and Cu(II)-to-Cu(I) Pro complexes by reducing agents that can promote the initial step in the Fenton reactions, which later enhance the reactive HO • radical production [41,46,60]. The reducing agents are superoxide anion (O † -  Table 3. Gibbs free energies (Δ r G 0 ), nuclear reorganization (λ), Gibbs free energies of activation (ΔG ‡ ), the diffusion rate constants (k D ), thermal rate constants (k) and the apparent rate constants (k app ) of reduction reactions of Fe(III)-to-Fe(II) complexes by superoxide radical anion (O 2

Direct enhancement
In this section, we investigate the oxidation of the formed Fe(III)-and Cu(II)-Pro complexes by HO • radical and compare them with that of the free Pro. We will discuss the FHT reactions between the Table 4. Gibbs free energies (Δ r G 0 ), nuclear reorganization (λ), Gibbs free energies of activation (ΔG ‡ ), the diffusion rate constants (k D ), thermal rate constants (k), and the apparent rate constants (k app ) of reduction reactions of Cu(II)-to-Cu(I) complexes by superoxide radical anion (O † -2 ) and ascorbate anion (Asc -) at 298.15 K. Units of energy values and reaction constants are in kcal mol −1 and M −1 s −1 , respectively.  Figure 5 represents the optimized structures for transition states of the FHT reaction between HO • radical and the two Site-2 complexes of Fe(III)-Pro and Cu(II)-Pro at H10 and H12. The Cartesian coordinates, and thermochemistry data of these structures are displayed in electronic supplementary material, table S9 (ESI). Figure 6 shows almost no difference between the TSs structures at the interactive sites for the complexes and the free ligand. Indeed, the C-H distances vary from 1.16 to 1.17 Å, while those of the O-H are from 1.44 to 1.45 Å, and the interactive angles vary from 166.9 to 175.9°, which are almost identical to those in the TSs of free Pro (figure 2). Despite many attempts, the TS structures for the Fe(II)-Pro and Cu(I)-Pro complexes cannot be located. As reported in previous studies, the FHT reactions between these complexes and HO • radicals may likely transform into SET reactions with reaction rates close to the diffusion limit [61,62].
The kinetics of the reactions between Fe(III)-Pro and Cu(II)-Pro complexes with HO • are resumed in table 5. As can be seen, the Gibbs free energies (Δ r G 0 ) for the FHT reactions at the H10/H12 position are −23.6/−24.0kcal mol −1 and −23.3/−23.3 kcal mol −1 for Fe(III)-Pro and Cu(II)-Pro complexes, respectively. Furthermore, the rate constants are equal to 2.99 × 10 7 /7.10 × 10 6 M −1 s −1 and 2.76 × 10 7 / 1.84 × 10 7 M −1 s −1 , respectively. These values are similar to those of the free Pro (table 2). Similarly, the rate constants for SET reactions of the complexes are also as negligible as those of free Pro.
As reported above, the reactions of Fe(II)-and Cu(I)-Pro complexes may occur via only the single electron transfer (SET) process, with the apparent rate constants close to the diffusion limit being 2.03 × 10 9 and 6.69 × 10 9 M −1 s −1 for Fe(II)-Pro and Cu(I)-Pro complexes, respectively. Those values are slightly higher than the overall rate constant of the free Pro, i.e. 6.04 × 10 8 M −1 s −1 at 298.15 K (table 2). Overall, the complexation of Pro with the low oxidation state-based transition metal ions [Fe(II) and Cu(I)] enhances its oxidation rate by HO • radicals more straightforwardly and more spontaneously via the SET reaction. Conversely, we do not observe any enhancement for the Pro oxidation when this amino acid chelates the high oxidation state of transition metal ions [i.e. Fe(III) and Cu(II)].
Finally, we attempt to provide deeper insight into the chemical nature of the FHT reaction between the free Pro amino acid towards HO • radicals and the possible effect of the Fe(III) or Cu(II) ions complexation. We analyze in detail the NPA atomic charges and Hirshfeld spin densities for the H-acceptor (the O atom of HO • radical), the transferred-H (H10 and H12), and the H-donor (the C atom at the interactive site) along the intrinsic reaction coordinates (IRC) of the reactions between free Pro, Fe(III)-Pro, and Cu(II)-Pro complexes and the HO • at H10 and H12 sites. The singly occupied molecular orbitals (SOMO) and the SOMO-1 distributions for the transition states of the studied reactions are also evaluated. Hydrogen transfer, a critical step in many chemical, environmental and biological processes, is one of the fundamental chemical reactions. This process involves the transfer of two elementary particles, a proton and an electron, simultaneously or sequentially by hydrogen atom transfer (HAT) or proton-coupled electron transfer (PCET) process, respectively [63][64][65][66]. Distinguishing the chemical nature of these two processes has been a big challenge. In principle, HAT reactions which are adiabatic processes consist of the simultaneous transfer of an electron and proton from an H-donor to an H-acceptor without significant redistribution of molecular charge, corresponding to small solvent reorganization energies. Conversely, PCET reactions are non-adiabatic processes; thus, they typically involve different donors and acceptors for the electron and proton, and they are associated with significant molecular charge redistribution, tending to more powerful solvent reorganization energies. As a result, the electronic wave function of HAT reactions varies smoothly with proton coordination, whereas PCET reactions change suddenly and sharply [67]. Understanding these processes is crucial in evaluating critical biological reactions such as respiration, nitrogen fixation, photosynthesis and energy conversion in  royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 10: 230114 artificial photosynthesis or fuel cells [68]. Thus, in the final section of this paper, we contribute our attempts to answer whether the formal hydrogen transfer (FHT) between the free Pro amino acid with HO • radicals occurs via HAT or PCET mechanisms, and the complexation towards the transition metal ions change the chemical nature of these mechanisms. Figure 6 illustrates the evolution of NPA atomic charges and Hirshfeld spin densities for the H-acceptor (the oxygen atom of HO • radical), shifted-H (H10 and H12) and H-donor (the carbon atom at the interactive site) along the intrinsic reaction coordinates (IRC) of the reactions between free Pro, Fe(III)-Pro and Cu(II)-Pro complexes with HO • at H10 and H12 sites in the aqueous phase.
Regardless of the reaction of Pro at H10, the spin densities of the shifted-hydrogen (shifted-H) are relatively high, approximately 2.7 × 10 −2 at the IRC point being 0.0 (TS point), and increase to values of about 6.2 × 10 −2 at the product complex (PC) (figure 6a). These values correspond to hydrogen radicals transferred during a HAT reaction [64]. Furthermore, the spin density of the oxygen atom of HO  Table 5. The Gibbs free energies (Δ r G 0 , kcal mol −1 ), Gibbs free energies of activation (ΔG ‡ , kcal mol −1 ), diffusion rate constants (k D , M −1 s −1 ), thermal rate constants (k, M −1 s −1 ), apparent rate constants (k app , M −1 s −1 ) and branching ratios (Γ, %) of the FHT at H10 and H12, and SET reactions between the Fe and Cu complexes at Site-2 with HO • radical calculated in water at 298.15 K. royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 10: 230114 Finally, the singly occupied molecular orbitals (SOMO) and SOMO-1 distributions for the transition states are evaluated and displayed in figure 7. The SOMO orbitals at the interactive sites lie on the AH····H···DH vectors and displays node plane at the shifted-H position, which has the SOMO orbital characteristics of the HAT transition state [63][64][65]. Moreover, for the free Pro and Cu(II)-Pro oxidation, the hydro transfer happens without the participation of the HOMO orbital (figure 7a and c). However, the Fe(III)-Pro and HO • radical reaction is contributed by both SOMO and SOMO-1 orbitals. In addition, the SOMO and SOMO-1 energies vary from −10.06 to −9.93 eV and −11.02 to −10.56 eV, respectively, which are relatively lower than free Pro, from −9.48 to −9.22 eV and −9.51 to −9.50 eV, in turn. This observation explains why the oxidations of these complexes by HO • are more difficult than free Pro, which is in good agreement with the kinetic results (table 5).
In summary, the oxidations of Pro and its complexes by HO • radical occur via the HAT mechanism. In addition, the HAT reactions between Pro by HO • become more complicated when Pro chelates transition metal ions, including Fe(III) and Cu(II).

Conclusion
The oxidation of L-proline (Pro) by HO • radicals, as well as the influence of its complexation with Fe and Cu ions on this process, have been studied in the aqueous phase using the density functional theory (DFT) approach. The conclusions are multiple as follows: -FHT reactions of dipole-salt form play a crucial role in the oxidation of Pro by HO • radical, which occurs at various sites with the highest branching ratio found at H10 (30.04%) and H12 (25.60%). The total branching ratios for FHT reactions at C5 and C6 carbons are more predominant than those at C4 and C7. The overall rate constant is 6.04 × 10 8  royalsocietypublishing.org/journal/rsos R. Soc. Open Sci. 10: 230114

13
-The indirect effect of transition metal ions complexation on the oxidation of Pro is complicated depending on the chemical nature of the metal ions. The Fe(III)-Pro complexes do not induce any remarkable risk of reactive HO • radical formation. By contrast, the complexation of Pro toward Cu(II) ions enhances the appearance of the HO • radicals via the Cu(II)-to-Cu(I) Pro complex formation in the presence of O † -2 and Asc − reducing agents. -The direct effect of the chelation of Pro towards transition metal ions is evaluated through the reactions between the most stable complexes of Pro (i.e. Fe-Pro and Cu-Pro at Site-2) with HO • radical. As a result, the FHT reactions of the high oxidation state Fe(III) and Cu(II) complexes at H10 and H12 sites occur at slower rate constants than those of the free Pro. Meanwhile, the rate constants of the SET reaction of these complexes are negligible. Conversely, the rate constants of the low oxidation state-based ion complexes (i.e. Fe(II) and Cu(I)) are determined to be higher than those of the free ligand. -The HO • -initiated oxidation of Pro occurs via the hydrogen atom transfer (HAT) process. The complexation of free Pro towards Fe(III) and Cu(II) ions does not change the nature of their HAT processes.
Hopefully, the obtained results shed more light on the chemical mechanism of the L-proline amino acid oxidation by HO • radical and the effects of the complexation with the transition metal ions on the oxidation process. They contribute to a better understanding of the origin of ageing and different diseases resulting from the accumulation of carbonyl derivatives caused by protein oxidation.
Data accessibility. All relevant necessary data to reproduce all results in the paper are within the main text, electronic supplementary material [69] and the Dryad Digital Repository: https://doi.org/10.5061/dryad.k6djh9wbb [70].